Caloric Restriction (CR) is the most potent, robust, and reproducible known means of extending longevity and decreasing morbidity in lab rodents. Despite 70 years of research, the relevance of this observation for humans remains unknown. Relevance is supported by the established link between obesity and morbidity in humans. Potential linkages are being directly addressed by the NIH-sponsored CALERIE study. This study has enrolled ~220 individuals and collected blood samples at 6 time points over 24 months. Our proposed ancillary study has two goals: (A) to support CALERIE by characterizing the plasma lipidome in these samples by using a newly developed, high resolution liquid chromatography-mass spectrometry-based profiling approach that enables both qualitative and quantitative detection of >425 structurally identified individual lipids in biological samples (e.g, >100 unique triglycerides), and; (ii) to link these data with our existing metabolomics profiling data within the CALERIE cohort and with an equivalent study in rats fed ad libitum or with a series of diets varying in the extent and duration of caloric restriction. The long-range goals of these latter studies are to develop diet-based biomarkers that have utility in human epidemiological studies (for objective recognition of diet) and which predict disease risk in humans (e.g., breast cancer). In support of this proposal, we have shown that CR induces changes in blood lipids in rats, e.g., a reduction in overall circulating triglycerides. Our finer resolution enables us to also show that there are multiple lipid species specific changes, and that these changes differ in both direction and magnitude, even within a single class of lipids such as triglycerides. Furthermore, we have conducted a pilot lipidomics study of ~400 samples drawn from CALERIE, and we show that the signal we can follow is so strong that, even unblinded (and thus including mixed controls and restricted, at a 1:2 ratio), we are able to show the effect on blood lipids with the CALERIE protocol, and we are able to show similar changes in rats and humans. We hypothesize that the proposed high resolution lipidomics study will reveal additional benefits of low calorie diets in humans. The Aims are: Aim 1: To develop, optimize, and validate a defined series of nested plasma lipidomics-based biomarker profiles Aim 2: To determine the similarities, differences, and interactions between the systemic lipidomics profiles and systemic metabolomics patterns Aim 3: To determine the similarities, differences, and interactions between the systemic lipidomics profiles in humans and those in CR animals The proposed study furthers NIH goals of focusing on health and early interventions rather than late stage disease, and is well within our lab's capacity to complete in 2 years.